SOLUBLE LIPOXYGENASE ISOFORMS FROM TOMATO FRUIT

Extraction of maximum lipoxygenase activity from tomato fruits was sho wn to require the presence of 0.1% Triton X-100. The detergent appeare d to have a dual role improving recovery and preventing enzyme inactiv ation, especially during mechanical homogenization. Ultracentrifugatio n of total lipoxygenase activity revealed that the majority (96%) was of a soluble form, with very little associated with the membrane fract ion. By ammonium sulphate fractionation and anion-exchange chromatogra phy, soluble lipoxygenase was purified 46-fold. Separation of lipoxyge nase activity by isoelectricfocusing and detection by in-gel activity staining resulted in the identification of two predominant isoforms wi th pi values of 4.8 and 5.0, one of which appears to be ripening-speci fic. On examination of reaction products by HPLC, both isoforms appear ed-to exclusively produce 9-hydroperoxides. (C) 1997 Elsevier Science Ltd. All rights reserved.